Drug addiction is caused by the long-lasting effects of drugs of abuse on neural circuits in the brain. Initial, acute exposure to drugs is followed by transition of some users to chronic use, a subset of which experience loss of control over drug consumption and develop addiction. Drugs alter the endogenous reward circuitry in the brain to cause increased assignment of incentive salience to drug cues, effectively making drug-associated environmental stimuli more difficult to ignore and leading to intense craving and relapse to drug seeking. Propensity to relapse persists in abstinent addicts even long after withdrawal symptoms have subsided, demonstrating the establishment of a pervasive memory.The nucleus accumbens (NAc), a central integrator of the brains’ reward circuitry, has been extensively associated with the development of addiction. Defining the architecture and logic of the gene regulatory networks underlying the development of the behavioural response to drug experience, and specifically, understanding the mechanisms underlying the rewiring of transcriptional programs that occurs following chronic drug exposure, will provide insight into the development of addiction, as well as novel targets for therapeutic intervention.

In the period since the beginning of the project, we have successfully established a productive research lab, located in a newly renovated space (as of July 2013) and equipped with all the necessary equipment for implementation of the research plan proposed in the grant application. The lab currently comprises of a PhD-level research associate and lab manager, a postdoctoral scholar, 8 graduate students (4 PhD and 4 MSc) and 4 undergraduate trainees. The essential experimental setups have been developed, including establishment of a dissociated neuronal culture system, and the necessary setups for viral vector cloning, virus production, stereotactic surgery, behavior (behavioral sensitization and conditioned-place preference), fluorescently-directed dissection of virally-infected brain structures, and high-throughput analysis of transcription by microfluidic qPCR and RNAseq.

Using the experimental setups in the lab, we have replicated our preliminary findings, both for neuronal cultures, as well as for in-vivo transcriptional rewiring following behavioral sensitization to cocaine. We have also established shRNA constructs for relevant genes and are working to develop CRISPR gRNAs to target genes of interest, as a complementary approach. However, we are yet to investigate their function on the transcriptional response to stimulation, or on the rewiring of transcription following chronic cocaine experience.

An unexpected and highly promising avenue of research has developed from this work, which promises to expand to a number of new avenues of research. Studying the transcriptional programs induced by cocaine in naïve mice, or mice with different levels of prior exposure to the drug, we find induction of robust and highly reproducible transcriptional programs. This suggested to us that defined salient experiences may be encoded by unique patterns of transcription in relevant brain regions. We addressed this notion by careful development of 15 discrete salient experiences (such as development of a contingent sucrose habit, aversive experience of LiCl injections or footshock), studying the transcription programs induced in 8 different structures within the mouse brain (prefrontal cortex, anterior cingulate cortex, nucleus accumbens, dorsal striatum, amygdala, lateral hypothalamus, hippocampus and ventral tegmental area) using high-throughput microfluidic qPCR and RNAseq.The result of this study is the observation of very low variation between individual mice in the transcriptional response to a given experience, whereas each experience had a very robust and reliable transcriptional signature associated with it, in relevant brain regions. The transcriptional patterns we observed were clear enough to enable the decoding of the recent experience of each individual mouse with above 90% accuracy. Furthermore, a small subset of the transcriptional response is sufficient for this decoding.We believe this new approach to the investigation of experience-dependent plasticity, which we coin “behavioral transcriptomics” is a widely applicable approach to investigating the encoding of experience in the brain, as well as the formation of habits and compulsions. We are currently writing up this work for publication (aiming at submission to the journal ‘Nature’) and are hopeful this approach could form the basis for a new field of research in neuroscience.

The expected final results of the project will be a deeper understanding of the mechanisms underlying the encoding of the long-term memory of cocaine experience in the brain, likely underlying the development of cocaine addiction. The initial avenues of research are expected to develop according to the original plan, in parallel to the development of new approaches.